Influenza A viruses (IAV) are significant human pathogens causing yearly epidemics and occasional pandemics. Past pandemics have resulted in significant morbidity and mortality. The 1918 influenza pandemic was thought to have resulted in the death of at least 675,000 people in the U.S. and 40 million people worldwide. Pandemics in 1957 and 1968, while less severe, were also of major public health importance. A novel influenza A virus of swine origin became pandemic in 2009, causing the first pandemic in 41 years. In addition, annual epidemic influenza cases are also very significant resulting in up to 49,000 deaths in the U.S. annually. Clinical natural history studies to compare natural influenza virus infections in immunocompromised, non-immunocompromised, and other high-risk populations, including pregnant women, are important to better treat and prevent influenza virus-infected patients. We plan to ultimately recruit up to 1000 patients with influenza virus infection. Recruitment occurred in both inpatient and outpatient settings at multiple sites including the NIH Clinical Center, Suburban Hospital and the Washington Hospital Center. Careful clinical evaluation, analysis of viruses collected from patients, and studies of the immune response of the patients is being performed. An ongoing effort is continuing to analyze the data collected over the past 4 years of this study and a report on these data is planned within the next 6 months. As an adjunct to the natural history studies, the first human volunteer influenza virus challenge study to be performed in the U.S. in over a decade began this year using a 2009 H1N1 virus under an FDA-approved IND. A healthy volunteer screening study began at the Clinical Center to identify patients who will qualify and be available for these challenge studies in the upcoming months. The initial challenge study protocol, a dose finding study, is currently underway. Additional viral seed stocks of recent H3N2 and seasonal H1N1 viruses are being prepared. Since the emergence of the 2009 pandemic H1N1 virus, reports have identified an increased prevalence of the hemagglutinin receptor binding domain D222G mutation in viruses isolated from individuals who have suffered such severe infections, but this association is still unclear. Virus isolated from a nasopharyngeal wash of a severely ill immunocompromised patient at the time of diagnosis contained the D222, but isolates collected later in his course from a bronchoalveolar lavage contained primarily the G222 mutation and was mixed with a minor population of D222. These clinical isolates were compared to a G222 plaque purified virus in the ferret model. The G222 predominant clinical isolate was the most pathogenic in ferrets and developed the most diversity at the 222 amino acid position during infection, suggesting that increased diversity and not a specific polymorphism at HA 222 may be important in predicting pathogenic potential.